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5 protocols using α methyl dl tyrosine

1

Pharmacological Evaluation of PV3 in Rodents

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All drugs were diluted in sterile saline (0.9% NaCl). The drugs tested were: i) PV3 at doses of 5, 50, 100, 500, and 5000 µg/kg; ii) the benzodiazepine diazepam (Sigma, USA) at 2 mg/kg, used as positive control for anxiety and locomotion tests; iii) the tricyclic antidepressant imipramine (Sigma) at 15 mg/kg used as positive control in depression tests; and iv) the inhibitor of tyrosine hydroxylase α-methyl-DL-tyrosine (AMPT) (Sigma) at 200 mg/kg, used to reveal the contribution of catecholaminergic pathways. All drugs and vehicle were injected by the ip route.
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2

Manipulating Dopamine Signaling in Drosophila

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To influence dopamine signaling, 2 days old radish1 or CS flies were put for 14h on 5mg of Methylphenidate hydrochloride (Sigma) mixed under 10ml of regular food. To inhibit dDAT or DopR1 function, we kept 2 days old CS flies for 14h on 10ml of regular food with 30mg Desipramine hydrochloride or 1mg (+)-Butaclamol hydrochloride (Sigma), respectively. For inhibition of dopamine synthesis, freshly hatched CS flies were put on 20ml of regular food with 8mg α-Methyl-DL-tyrosine (Sigma) for 120h. We verified uptake of food by the addition of a non-hazardous blue dye, which after 14h stained the abdomen of the flies. To ablate the mushroom bodies, CS flies were treated with Hydroxyurea (Sigma) [16 (link)]. Dopamine was measured by HPLC (M. Krischke, Botany Department, University of Wuerzburg).
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3

Pharmacological Modulation of Neuronal Signaling

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Picrotoxin and MK-801 were from Abcam. CGP55845, DNQX, Dihydro-β-erythroidine hydrobromide, SCH 23390 hydrochloride, and sulpiride were obtained from Tocris Bioscience. BAPTA was obtained from Invitrogen. ω-Conotoxin GVIA and ω-Agatoxin TK were from Alomone Labs. All the other chemicals (α-methyl-DL-tyrosine, DEAB, TTX, 4-Aminopyridine, EGTA-AM, BAPTA-AM, scopolamine hydrobromide) were from Sigma-Aldrich.
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4

Measuring Catecholamine Turnover Dynamics

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Catecholamine turnover was measured on the basis of the decline in tissue NE content after the inhibition of catecholamine biosynthesis with α-methyl-dl-tyrosine (α-MPT) (200 mg/kg i.p. injection; Sigma-Aldrich, ST, Quentin, France), as described previously (44 (link)).
In the morning, bedding was changed, and C57BL/6N mice were food-deprived for 3 hours to insure postprandial state and injected with α-methyl-dl-tyrosine (α-MPT; a tyrosine hydroxylase inhibitor) to block catecholamine synthesis. Before (time = 0) and 3 hours after the injection (time = 3 hours), animals were euthanized, and the tissues were removed, flash-frozen in liquid nitrogen, and stored at −80°C for monoamine and metabolite analysis.
Catecholamine content at time = 0 [NE (0)] was determined on a group of animals receiving a saline injection. Because the concentration of catecholamine in tissues declined exponentially, we could obtain the rate constant of NE efflux (expressed in h − 1). Comprehensive analysis of NE was carried out by reverse-phase liquid chromatography (LC) with electrochemical detection as described in (67 ). The values obtained were expressed as nanogram per milligram wet tissue and were logarithmically transformed for calculation of linearity of regression, SE of the regression coefficients, and significance of differences between regression coefficients.
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5

Pharmacological Modulation of Neuronal Signaling

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Picrotoxin and MK-801 were from Abcam. CGP55845, DNQX, Dihydro-β-erythroidine hydrobromide, SCH 23390 hydrochloride, and sulpiride were obtained from Tocris Bioscience. BAPTA was obtained from Invitrogen. ω-Conotoxin GVIA and ω-Agatoxin TK were from Alomone Labs. All the other chemicals (α-methyl-DL-tyrosine, DEAB, TTX, 4-Aminopyridine, EGTA-AM, BAPTA-AM, scopolamine hydrobromide) were from Sigma-Aldrich.
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